Direct use of tobacco products increases an individual's susceptibility to cancer. Nicotine-derived nitrosamines are cancer suspect agents that are found in tobacco, smokeless tobacco, tobacco smoke, and sidestream (environmental or secondary) smoke. The pervasive nature of these compounds has generated interest in their role as causative factors in tobacco-related malignancies. Although much work has addressed the carcinogenic nature of the nicotine-derived nitrosamines, little work has appeared in the literature connecting adducts formed by these compounds to tumorigenesis at the molecular level. Knowledge concerning DNA damage and repair following exposure to carcinogens is critical to understanding the persistence of adducts in DNA, and as a consequence, the formation of mutations leading to neoplasia. The compound 4-(methylnitrosamino)-l-(3-pyridyl)-l-butanone (NNK) is considered the most carcinogenic of the nicotine-derived nitrosamines. We will map the adducts formed by NNK that are repaired by base excision repair, nucleotide excision repair, and O6-methyguanine-DNA methyltransferase at nucleotide resolution in human cells in two genes closely associated with tumorigenesis: P53 and Ki-Ras. Ultimately, we will compare damage and repair rates in our experiments to the mutations found in those genes in tobacco-related tumors. We will also determine the contribution of different DNA repair systems to the eradication of adducts formed by NNK. The goal of this project is to establish the role of NNK in the etiology of tobacco-related, tobacco smoke-related, and secondary tobacco smoke-related cancers.